1. Field of the Invention
The present invention relates to a microparticulate diet for the culture of larval fish and crustaceans. In particular, the present invention relates to a proteinaceous and nutritionally complete formulated food product to replace live feed as the diet for larval fish and crustaceans using Macrobrachium rosenbergii as the test organism.
2. Discussion of the Background
The availability and nutritional value of live food is considered to be a limiting factor in larviculture of many fish and crustacean species. Larvae of many species have been successfully raised using live food (e.g., Artemia nauplii and rotifers). However, there are several major disadvantages that are associated with the use of live food, such as variable nutrient composition, the potential for introduction of a pathogen into the culture system, and the amount of labor required for preparation of the live feed. Formulated feed is an attractive and valuable alternative to live food. For example, successful formulated diets can be used in nutritional studies and can serve as a vehicle for the delivery of hormones and therapeutic agents. Moreover, formulated diets introduce flexibility in adjustment of size.
Newly hatched Artemia nauplii and juveniles are commonly used as food for larval culture. However, the results of past research have led to concern about lack of consistency in the nutritional value of Artemia. For example, the nutritional quality and physical properties of Artemia vary depending upon the source and time of harvest of cysts (dormant eggs). In response to potential nutritional deficiencies, Artemia are commonly enriched with HUFA (highly unsaturated fatty acids) and other nutrients before feeding them to larval fish and crustaceans. (Clawson and Lovell, 1992). This enrichment is achieved by feeding algae, enriched yeast cells, or encapsulated, emulsified lipids to the larvae. The availability of Artemia cysts is subject to change. Because the availability and nutrient quality of Artemia is inconsistent, there is a need for a complete, formulated feed.
Numerous attempts to develop a formulated diet for larvae of different species have been made, but none have resulted in an effective replacement for live food. For example, microencapsulated, microbound, and microcoated formulated diets have been developed but with limited success. A microencapsulated diet is prepared by encapsulating a nutrient solution, colloid or suspension within a natural or synthetic polymer. A microbound diet is prepared by binding ingredients (usually powdered) together with a binder such as agar, alginate, or carrageenan. A microbound diet coated with materials like wheat gluten or cholesterol is termed a microcoated diet. (Bautista et al, 1989). The data collected suggest that practical diets containing raw, unrefined ingredients appear to be more effective than semi-purified diets. However, despite significant effort, limited success has been realized due to a presumed low capacity of larvae to digest these diets and the virtually unknown nutritional requirements of the larvae. Some of the previous attempts to make a formulated diet are illustrated below.
1. Bautista et al. (1989) developed a kappa-carrageenan microbound freeze-dried diet for larval P. monodon. This diet was composed primarily of squid, shrimp meals and bread flour. A 55% survival from zoea to postlarvae was achieved with this diet.
2. Villamar et al. (1993) created water-stable lipid wall microcapsules with gelled alginate-gelatin and evaluated the ability of larval Penaeus vannamei to digest the microcapsules by measuring accumulation of 14C. Minimum ingestion rates of the microcapsules were determined to be 48-99 xcexcg dry weight of larva/day.
3. Jones et al. (1987) designed microencapsulated diets for several species of penaeid shrimp. However, higher survival was achieved when larvae were fed free particulates rather than microencapsulated diets (68 vs. 50%). In a later study, Kumlu and Jones (1995) reported slower growth and lower survival of P. indicus larvae fed microencapsulated diets unless an algal supplement was provided as part of the diet.
4. Koshio et al. (1989) reported good growth and survival of larvae of the prawn Penaeus japonicus when a diet consisting of egg yolk powder, casein, albumin, amino acid mix, soybean lecithin, cholesterol, vitamin mix, mineral mix, glucosamine chloride, and alpha cellulose was fed. The total protein content of this diet was 48.2%.
5. Ohs (1995) developed semi-purified spray-dried diets and evaluated their performance with larval striped bass, Morone saxatilis, and a freshwater prawn. Although the larvae of both species consumed the diets, growth and survival were significantly less than that of Artemia-fed larvae. Wan (1999) developed several semi-purified microparticulate diets and studied the influence of the weaning period and supplementation of live food on performance. The survival and growth of larvae of M. saxatilis and M. rosenbergii fed this microparticulate diet remained inferior to that achieved when fed Artemia nauplii. 
More recent studies have shown that the contribution of the autolytic enzymes from consumed prey contribute minimally to digestion in the larval gut. (Garcia-Ortega et al., 1998 and Sorgeloos, 1999). Live food can be, at least theoretically, completely replaced by an appropriately prepared formulated diet. To ensure long term survival and growth of the larvae, the diet should be highly attractive, digestible, have a comparable nutrient composition to that of live food, and have adequate physical properties such as water stability and neutral buoyancy. (Bengston, 1993.)
Use of inert feed, such as fish or mollusk flesh or farm-made feeds that contain fish or mollusk flesh are almost exclusively used as supplements. (Lavens et al., 2000). Although currently used formulated feed are effective alternative nutrient sources and contribute to a reduction in operational costs, they do not address the critical need for control and consistency of nutrient composition. In addition, their contribution to the overall satisfaction of nutritional requirements cannot be discriminated.
The culture of many cultivated species, e.g., freshwater prawn (Macrobrachium rosenbergii) and striped bass (Morone saxatilis), is limited by the high cost of juveniles. For example, up to 60% of the cost of these species is associated with the cost of feed. Thus, the development of a comparatively inexpensive formulated diet capable of partially or completely replacing live food will not only decrease production costs associated with aquaculture enterprise but will also provide a foundation for further nutritional studies. Furthermore, successful formulated diets will help to make larviculture a more predictable and cost-effective by reducing potential sources of larval diseases, simplifying hatchery design, and providing a reliable supply of a nutritionally balanced food that is always available and is not subject to a change in the nutrient composition. Thus, the use of formulated diets for larval culture should contribute to the development of a sustainable aquaculture enterprise throughout the world through lower production costs and greater availability of juveniles of commercially cultured species of fish and crustaceans.
It is an object of the present invention to provide a formulated microbound diet for the culture of larval fish and crustaceans.
It is another object of the present invention to provide a nutritionally complete formulated diet.
It is a further object of the present invention to provide a cost-effective formulated diet.
It is yet another object of the present invention to provide a particulate food product for consumption by larval fish and crustaceans.
It is another object of the present invention to provide a formulated food product that is alginate-bound.
It is an advantage of the present invention that the formulated diet is less expensive than live feed such as Artemia nauplii. 
It is another advantage of the present invention that the formulated artificial diet is water stable.
It is another advantage of the present invention that the formulated diet can be prepared in either a moist or dry form.